Last‐Step Enzymatic [¹⁸F]‐Fluorination of Cysteine‐Tethered RGD Peptides Using Modified Barbas Linkers

Abstract

Abstract:We report a last-step fluorinase catalyzed [ 18 F]-fluorination of a cysteine-containing RGD peptide. The peptide was attached through sulfur to a modified and more hydrophilic variant of the recently disclosed Barbas linker which was itself linked to a chloroadenosine moiety via a PEGylated chain. The fluorinase was able to use this construct as a substrate for a transhalogenation reaction to generate [ 18 F]-radiolabelled RGD peptides, which retained high affinity to cancer cell relevant α v β 3 integrins

Enzymatic Fluorination of Biotin and Tetrazine Conjugates for Pretargeting Approaches to Positron Emission Tomography Imaging

“…Enzymatic transhalogenation in the presence of [ 18 F]fluoride was investigated with 16 : its conversion into [ 18 F]FDA‐PEG‐tetrazine [ 18 F] 17 was clearly demonstrated against a product reference. By using the previously established protocol, [ 18 F] 17 was generated with approximately 10 % radiochemical conversion (decay uncorrected) after 30 min incubation with the fluorinase (Figure S46). Isolation of [ 18 F] 17 by semipreparative HPLC afforded the radiolabelled product with two minor radiolabelled impurities (Figure S47).…”

“…After 30 min incubation, the enzyme was cleanly precipitated by heat denaturation and the solution was removed after centrifugation, prior to purification by HPLC. During semipreparative HPLC the eluted peak corresponding to [ 18 F] 27 was collected (Figure , t R =9.7 min), diluted with water (50 mL) and loaded onto a C 18 reversed‐phase cartridge.…”

“… A) The native reaction catalysed by the fluorinase enzyme. B) Fluorinase‐mediated transhalogenation of C‐2‐modified substrates 6 and 7 illustrates previously achieved alkyne–azide click and Barbas linker/thiol‐ligation bioconjugation strategies.…”

“…In this way the fluorinase will catalyse “last‐step” aqueous [ 18 F]fluorination of 5′‐ClDA‐PEG‐peptides . To date, methodologies permitting bioconjugation of RGD peptides to the fluorinase‐binding 5′‐ClDA moiety either through click‐based chemistry (by use of azide‐functionalised peptides) or by using a modified Barbas bioconjugation strategy (through the use of cysteine‐containing peptides) have been developed (Scheme ). Recently, fluorinase has also become the target of engineering through directed evolution, in an effort to achieve higher levels of radiochemical conversion (RCC) and to broaden the structural diversity of substrates…”

Arylierungschemie für die Biokonjugation

“…MSBT wies eine hervorragende Selektivitätfürdie Reaktion mit Cystein-Thiolen auf;andere Protein-Nukleophile wie die in den Seitenketten von Serin, Ty rosin, Tr yptophan, Methionin, Histidin und Lysin zeigten keine Reaktion. [82] Xian und Furdui entwickelten ein neues heteroaromatisches Alkylsulfon, 4-(5-Methylsulfonyl- [1,2,3,4]tetrazol-1-yl)phenol (MSTP), das fürd ie selektive Blockade von Protein-Thiolen ohne eine Kreuzreaktivitätm it Sulfensäuren geeignet ist. [77] Die Markierungsreaktion erwies sich als proteinkompatibel, was durch die Blockade des Thiols im aktiven Zentrum von Glyceraldehyd-3-phosphat-Dehydrogenase( GAPDH) unter milden Bedingungen bestätigt werden konnte.…”